Biology subjects

Cochrane, A.

Publications and source records attributed to Cochrane, A..

6 recordsLinked to original sources

bioRxiv2026

A cell line model for the study of CD4-negative HIV-1 infection and latent virus reservoirs

Although controversial, limited publications support the notion that HIV-1 can infect CD4-negative cells. The objective of this study was to provide a comprehensive investigation of a universally available CD4-negative cell line model system that can be infected with X4 and R5 HIV-1 to generate integrated proviral DNA and serve to study latent viral reservoirs. The reason that HIV-1 infection of CD4-negative cells has become less investigated is due to a lack of a fully characterized model for the study of this unusual pathway. To address this critical need, human osteosarcoma (HOS) cells, engineered to express either CD4, CCR5 or CXCR4, and easily available from a commercial source were used. CD4 expression was examined using western immunoblot, flow cytometry, anti-CD4 blocking antibody and mRNA expression. Cells were infected with HIV-1 pseudo-enveloped viruses bearing either JR-FL (R5-tropic) or HXB2 (X4-tropic) envelopes, constructed on NL4-3 luciferase/GFP backbone. Infection was monitored by luciferase readout and visualized by GFP immunofluorescence. Raltegravir was used to inhibit integration, and AMD3100 and maraviroc used to block chemokine coreceptors, CXCR4 and CCR5, respectively. Productive versus latent infection was quantified by dual-fluorescence readouts using HI.fate.E. We confirmed that HOS cells lack CD4. HOS cells expressing only CCR5 or CXCR4 supported HIV-1 infection, although infection was significantly lower than in matched CD4-positive controls. Raltegravir treatment blocked proviral integration in all instances. Coreceptor antagonism and envelope-deficient viruses revealed that infection of CD4-negative CXCR4 cells remained CXCR4-dependent, whereas CD4-negative CCR5 cells showed evidence of CCR5-independent infection. Dual-reporter HI.fate.E assays indicated that CD4-negative cells could support both productive and latent infection. These studies establish a universally available cell line model for the study of CD4-negative HIV-1 infection. This cell line model will provide insight into the question of how CD4-negative cells can be infected with HIV-1 and whether CD4-negative cells can provide latent viral reservoirs in HIV/AIDS. Author summarySince the first description of HIV/AIDS in 1981 and the recognition that CD4 was a primary receptor for HIV-1 in 1983, a limited number of reports have suggested that cells lacking CD4 could be infected with HIV-1. These reports continued even when it was shown in 1996 that co-receptors, CXCR4 and CCR5, were also required for HIV-1 infection of CD4 T-helper cells. Indeed, crystallography studies showed that CD4 was required to interact with the HIV-1 envelope gp120 in order to cause conformational changes in the envelope to expose the binding motif for chemokine co-receptor engagement, required for additional conformational changes to expose the gp41 fusion protein, allowing for entry and infection. However, reports continued that cells lacking CD4 could be infected which raised questions as to how this can happen. To address this critical gap, we have identified a cell line, HOS, that is commercially available, having expression of CD4, CXCR4 and/or CCR5. Using these HOS cell lines, we have been able to confirm that HIV-1, either X4 or R5 enveloped viruses, can infect CD4-negative cells. We have also confirmed that infection is productive and allows for latent proviral integration. Our findings provide a system for further studies of the mechanism(s) of HIV-1 infection of CD4-negative cells using a consistent model and may aid in elucidating establishment of viral reservoirs.

pathology↗

bioRxiv2026

Hepatic HIF2α modulates extra-hepatic disease-associated phenotypes during metabolic dysfunction-associated steatotic liver disease

Metabolic dysfunction-associated steatotic liver disease (MASLD) afflicts more than one-third of adults globally, contributing significantly to an increased cardiovascular disease risk. Further, patients with severe liver disease experience muscle weakness (sarcopenic obesity) and fatigue. Hypoxia-inducible factor 2 (HIF2) accumulates in the livers of MASLD patients and has been implicated in disease progression. Here we sought to understand the role of hepatic HIF2 in mediating hepatic and extra-hepatic features of MASLD. Using a well-validated obese mouse model of MASLD, we investigated the impact of hepatocyte-specific HIF2 deletion (hHIF2-/-) on hepatic, cardiac and skeletal muscle metabolism, and cardiac function. Over 28 weeks, mice were exposed to a high-fat, high-fructose, high-cholesterol (GAN) diet, which induced obesity alongside hepatic steatosis, fibrosis and inflammation. In contrast to observations in lean mouse models of liver disease, hHIF2-/- did not protect against MASLD, despite greater hepatic NADH-supported mitochondrial respiration and higher intracellular sphingomyelin levels. Instead, in the hearts of GAN-fed mice, hHIF2-/- caused diacylglycerol accumulation independent of diet, accumulation of long-chain acyl-carnitines and exacerbation of ceramide accumulation. Langendorff-perfused hearts from hHIF2-/- mice showed systolic and diastolic dysfunction, including 24% lower left ventricular developed pressure and 34% lower maximal rate of relaxation (dP/dtmin). However, isolated hearts from hHIF2-/- mice were protected against MASLD-associated sympathetic dominance, determined using autonomic receptor agonist stimulation. Both GAN-feeding and hHIF2-/- were associated with lower lean mass (14% and 5.4% lower than respective controls), whilst hHIF2-/- enhanced OXPHOS-associated protein levels in gastrocnemius muscle. Overall, hHIF2-/- resulted in detrimental extra-hepatic effects, including myocardial lipid accumulation, impaired cardiac function, and loss of whole-body lean mass, with no apparent protection against MASLD disease progression.

physiology↗

bioRxiv2025

Sleep is associated with reduction of excitatory signaling in medial prefrontal cortex

Although many sleep medications enhance inhibitory signaling, it remains unclear whether inhibitory or excitatory neurotransmitters contribute to the natural transition from wakefulness to sleep in humans. Here, we show that changes in excitatory, rather than inhibitory, neurotransmitter levels are associated with this transition. Young, healthy participants underwent two nap sessions with polysomnography, during which glutamate and GABA concentrations in the medial prefrontal cortex were measured using magnetic resonance spectroscopy. Glutamate gradually decreased during deeper sleep stages compared to wakefulness in the second session, with better sleep quality. No such change occurred in the first session with poorer sleep, likely due to the first-night effect. Furthermore, reduced glutamate significantly mediated sleep-onset latency in both sessions. Conversely, GABA concentration did not change from wakefulness to sleep in either session. These findings provide the first evidence that reduced excitatory signaling is a key feature of natural good sleep onset in the human brain.

neuroscience↗

bioRxiv2025

Concurrent multimodal measurement of excitation and inhibition shows that EEG-based estimates are linked to the ratio of glutamate to GABA

Neural excitation/inhibition (E/I) ratio is dynamically regulated on multiple timescales. Adaptive changes in E/I ratio can support healthy development, learning, and cognition, while disordered E/I ratio has been implicated in neurodevelopmental disorders, neurodegenerative disorders, and states of impaired vigilance. There has been growing interest in inferring E/I ratio from efficient and noninvasive measurements such as electroencephalography (EEG), and several algorithms have been proposed to estimate E/I ratio from EEG. Despite promising results, there has been a lack of validation studies testing the underlying neurochemical changes leading to increased or decreased EEG-based E/I ratio. Here, using concurrent EEG and magnetic resonance spectroscopy (MRS) for over an hour, we assessed which algorithm of EEG-based E/I ratio best matched with MRS-based E/I ratio in humans of both sexes. The MRS-based E/I ratio was obtained by the ratio of glutamate concentration to GABA concentration. We applied 10 candidate indices of EEG-based E/I ratio using four approaches in several spontaneous frequency bands. Uniquely, we quantified the associations between the EEG-based E/I ratio and MRS-based E/I ratio separately for between-subjects and within-subjects variations. We found that each EEG-based E/I algorithm showed reliable and positive associations with MRS-based E/I, and especially EEG-based E/I ratio in alpha band with a criticality theory based approach showed the best association to the MRS-based E/I ratio. While these associations were evident for between-subjects comparisons, they were quite weak for within-subjects comparisons. These results suggest that EEG-based E/I algorithms are likely to reflect, at least in part, relative concentrations of glutamate and GABA.

neuroscience↗

bioRxiv2024

Exploiting the Achilles' Heel of Viral RNA Processing to Develop Novel Antivirals

Viruses continue to pose a significant health burden to the human population, and recent history has shown a concerning surge in viral threats. Treatment options for viral infections are limited, and viruses have proven adept at evolving resistance to many existing therapies, highlighting a significant vulnerability in our defenses. In response to this challenge, we explored the modulation of cellular RNA metabolic processes as an alternative paradigm to antiviral development. Many viruses depend on the host cells RNA splicing machinery, and small alterations to this host process results in catastrophic changes in viral protein production, ultimately inhibiting virus replication. Previously, the small molecule 5342191 was identified as a potent inhibitor of HIV-1 replication by altering viral RNA accumulation at doses that minimally affect host gene expression. In this report, we document 5342191 as a potent inhibitor of adenovirus, coronavirus, and influenza replication. In each case, 5342191-mediated reduction in virus replication was associated with altered viral RNA accumulation and loss of viral structural protein expression. Interestingly, while resistant viruses were rapidly isolated for compounds targeting either virus-encoded proteases or polymerases, we have not yet isolated 534219-resistant variants of coronavirus or influenza. As with HIV-1, 5342191s inhibition of coronaviruses and influenza is mediated through the activation of specific cell signaling networks, including GPCR and/or MAPK signaling pathways that ultimately affect SR kinase expression. Together, these studies highlight the therapeutic potential of compounds that target cellular processes essential for the replication of multiple viruses. Not only do these compounds hold promise as broad-spectrum antivirals, but they also offer the potential of greater resilience in combating viral infections.

microbiology↗

bioRxiv2023

Obesogenic diet in mice leads to inflammation and oxidative stress in the mother in association with sex-specific changes in fetal development, inflammatory markers and placental transcriptome

IntroductionObesity during pregnancy is related to adverse maternal and neonatal outcomes. Factors involved in these outcomes may include increased maternal insulin resistance, inflammation, oxidative stress and nutrient mishandling. The placenta is the primary determinant of fetal outcomes, and its function can be impacted by maternal obesity. The aim of this study in mice was to determine the effect of obesity on maternal lipid handling, inflammatory and redox state, and placental oxidative stress, inflammatory signaling, and gene expression relative to female and male fetal growth. MethodsFemale mice were fed control or obesogenic high-fat/high-sugar diet (HFHS) from 9 weeks prior to and during pregnancy. On day 18.5 of pregnancy, maternal plasma, and liver, placenta, and fetal serum were collected to examine the immune and redox state. The placental labyrinth zone (Lz) was dissected for RNA-sequencing analysis of gene expression changes. ResultsHFHS diet induced, in the dams, hepatic steatosis, oxidative stress (reduced catalase, elevated protein oxidation) and activation of pro-inflammatory pathways (p38-MAPK) along with imbalanced circulating cytokine concentrations (increased IL-6 and decreased IL-5 and IL-17A). HFHS fetuses were asymmetrically growth-restricted, showing sex-specific changes in circulating cytokines (GM-CSF, TNF-, IL-6 and IFN-{gamma}). Morphology of the placenta Lz was modified by a HFHS diet, in association with sex-specific alterations in the expression of genes and proteins implicated in oxidative stress, inflammation, and stress signaling. Placental gene expression changes were comparable to that seen in models of intrauterine inflammation and were related to a transcriptional network involving transcription factors, LYL1 and PLAG1. DiscussionThis study shows that fetal growth restriction with maternal obesity is related to elevated oxidative stress, inflammatory pathways, and sex-specific placental changes. Our data are important given the marked consequences and the rising rates of obesity worldwide.

physiology↗